2.2.8 研究 | 2021-多组学分析肺结核队列的记忆T细胞状态

刘小泽写于2021.5.25

速览

题目：Multimodally profiling memory T cells from a tuberculosis cohort identifies cell state associations with demographics, environment and disease
作者：哈佛医学院 Soumya Raychaudhuri 团队
首先感觉他们实验室主页做的很不错：https://immunogenomics.hms.harvard.edu/lab
链接：https://www.nature.com/articles/s41590-021-00933-1
发表在：Nature Immunology （2021-05-24）

数据是公开的：GSE158769 ，采用CITE-seq

Platforms    GPL11154    Illumina HiSeq 2000 (Homo sapiens)
Samples      GSM4810298    Memory_T_cell_CITE-seq

GSE158769_exprs_norm.tsv.gz    3.0 Gb    (ftp)(http)    TSV
GSE158769_exprs_raw.tsv.gz    747.0 Mb    (ftp)(http)    TSV
GSE158769_meta_data.txt.gz    19.8 Mb    (ftp)(http)    TXT

Genotype data：phs002025.v1.p1

分析代码：https://github.com/immunogenomics/TB_Tcell_CITEseq
还发开了一个网页：https://immunogenomics.io/tbru/

摘要

分析了来自秘鲁的259个肺结核（tuberculosis , TB)病人的500,089个记忆T细胞，得到31种细胞状态
在排除年龄、性别、季节、遗传等因素影响后，发现了一种多能型type 17 helper T (TH17) 细胞状态，在结核杆菌发展成肺结核病的过程中丰度和功能都有所下降

方法

数据准备

PBMC sample preparation
Flow cytometry of total PBMCs
CITE-seq of isolated memory T cells
Bulk RNA sequencing
Genotyping and genetic data processing

数据分析

scRNA比对+定量
scRNA数据拆分
scRNA数据质控
降维
聚类+注释
bulk RNA-seq 比对+定量
利用MASC(modeling of associations of single cells )评价不同细胞状态与疾病的关系

先用CITE-seq测了>500,000个T细胞

在秘鲁首都利马进行了一次大型的流行病学调查（n= 18,544），从其中招募了264个人，采样了外周血单核细胞进行CITE-seq测定，同时还加入了基因表型数据和蛋白数据
图b是进行了6次QC，每一次过滤掉一些细胞，最后得到了500K
大部分样本的细胞数量在2k左右

得到记忆T细胞的31种不同状态

利用canonical correlation analysis (CCA) 进行降维，选择top20 canonical variates (CVs)进行批次校正和聚类，根据基因和蛋白marker得到31个细胞状态

23/31 were CD4+; five were CD8+; one (C-24) was a mixture; Two clusters (C-30 and C-31) were CD4−CD8−
C-20： contained a subset of CD4−CD8− and CD8+ cells expressing innate-like T cell markers, including ZBTB16 and CD161 and CD26 surface proteins
one central memory cluster (C-25) and distinct GZMK+ (C-28) and GZMB+ (C-29) effector subsets, reflecting different cytotoxicities
high expression of HLA-DR and CD38 surface protein and proliferation-associated MKI67 (C-15 and C-27) represent chronically activated cells
TH17 cells (C-12, CCR6+ and RORC)
type 1 helper T cells (TH1; C-17, CXCR3+ and IFNG and TBX21)
heterogeneous continuum of intermediate TH1/TH17 states (C-13, C-16 and C-19) with varying degrees of CXCR3, CCR6, CCR5 and CD161 surface protein expression and RORC and TBX21 expression
CD161+ subset of type 2 helper T (TH2) cells (C-14), described as pathogenic, with higher expression of allergy-associated HPGDS and IL17RB
a subset of FOXP3+ Treg cells (C-5) expressing higher CCR6 surface protein and CTLA4 and RORC than other Treg cells